In order to prepare antioxidant peptide through hydrolyzing low-value protein resources with bacterial extracellular proteases and to discover novel proteases, crude extracellular protease from Pseudoalteromonas sp. SHK1-2 was obtained through fermentation which was used to hydrolyze collagen extracted from Cirrhinus molitorella skin. Small peptide fraction was isolated from hydrolysate by ultrafiltration and Sephadex LH-20 size exclusion chromatography and showed 1, 1-diphenyl-2-picrylhydrazyl radical scavenging activity (35.6%±7%), oxygen radical absorbance capacity and inhibition of DNA oxidation damage. The molecule weight was 776.2 Da, and amino acid sequence was Thr-Ala-Gly-His-Pro- Gly-Thr-His through liquid chromatography mass spectrum. Our findings suggest that peptide obtained from low-value protein of fish waste by hydrolysis with bacterial protease has antioxidant activity.
为了以细菌胞外蛋白酶酶解低值蛋白资源制备抗氧化活性肽以及挖掘新型蛋白酶，采用液体发酵培养的方法对假交替单胞菌Pseudoalteromonas sp. SHK1-2 进行发酵产酶，获得胞外蛋白酶粗酶液用于酶解热水法提取的鲮鱼胶原蛋白，通过超滤、sephadex LH-20 分子筛层析获得具有DPPH 自由基清除能力 (35.6%±7%)、氧自由基清除能力 (Oxygen radical absorbance capacity，ORAC) 及DNA 氧化损伤抑制活性的小分子肽，液相色谱质谱联用鉴定该活性肽分子量为776.2 Da，预测氨基酸序列为Thr-Ala-Gly-His-Pro-Gly-Thr-His。ORAC检测验证了人工合成的多肽同样具有抗氧化活性。研究表明细菌胞外蛋白酶在低值资源高值化中具有一定的应用前景，对于新型蛋白酶及其应用的挖掘具有一定的参考意义。.
Keywords: Corrhinus molitorella; DNA oxidation damage; DPPH; ORAC; antioxidant peptide; collagen.